O-GlcNAc addition to serines and threonines of cytosolic and nuclear proteins is a unique carbohydrate modification with no known function. This proposal addresses the potential for O-GlcNAc to be a dynamic switch mechanism controlling the functional states of proteins (analogous to regulated phosphorylation in signal transduction). In the context of signaling, the role of O-GlcNAc in regulating various functional aspects of Raf-1 will be examined. The potential for O-GlcNAc to regulate kinase activity or interaction of Raf-1 with key proteins such as Ras and 14-3-3 in a basal or inducible way will be studied.